The present invention relates to the field of cell proliferation-inhibiting proteins. More particularly, the invention relates to a protein comprising or consisting of a VPg protein, fragments thereof or analogs thereof, that are cytotoxic to eukaryotic cells, and/or inhibit the cell proliferation. The invention also relates to medical uses of such a protein, fragments thereof or analogs thereof.
Eukaryotic initiation factor eIF4E is a mRNA 5′ cap-binding protein, which both attaches mRNA and binds the eIF4G component of the eIF4F initiation complex. A significant fraction of eIF4E (up to nearly 70%) localizes to the nucleus (Lejbkowicz et al., 1992). It has been shown that nuclear eIF4E is involved in the nucleocytoplasmic transport of certain messenger RNAs (Lejbkowicz et al., 1992; Rosenwald et al., 1995; Rousseau et al., 1996; Lai and Borden, 2000). It has been demonstrated that the unchecked over-expression of eIF4E can cause cell growth and malignant transformation, whereas reduction of the eIF4E level can reverse the transformed phenotype (reviewed in Sonenberg and Gingras, 1998). Furthermore, in humans, some tumor types exhibit elevated levels of eIF4E (Ruggero and Pandolfi, 2003). It appears that at high eIF4E levels there is increased cytoplasmic level of proteins involved in the malignant transformation, due to the increase in the eIF4E-dependent transport of the appropriate mRNAs to the cytoplasm (Shantz and Pegg, 1994; Rousseau et al., 1996; Lai and Borden, 2000).
International Application No. WO 00/78803 describes eIF4E- binding peptides useful in therapy, and particularly for the induction of programmed cell death, presumably by disrupting the formation of the eIF4F complex. All these peptides comprise the amino acid sequence (motif) YxxxxLØ(SEQ ID NO: 31), wherein x is any amino acid and Ø is Leu, Met or Phe, required for binding human eIF4E (accession number NP—001959 in the GENBANK database, Apr. 15, 2007 version).
Thus, there is an interest in the development of adequate eIF4E binding agents for use in anti-cancer therapy. There is also a need to find eIF4E binding agents possessing a distinct mechanism of action from that described above, to overcome drug resistances for instance.
Potato virus Y (PVY) is the type member of the Potyvirus genus, belonging to the Potyviridae family, the largest and economically most important group of plant viruses. Potyviruses have a so-called genome-linked VPg protein (hereinafter referred to as VPg protein or VPg) covalently attached to the 5′ end of their linear nucleic acid genomes (Riechmann et al., 1989 and Murphy et al., 1991). The potyvirus VPg protein is implicated in virus protein synthesis, long-distance movement in plant tissue and virus replication (Lellis et al., 2002; Schaad et al., 2002; Fellers et al., 1998). Early after potyvirus infection, the genomic RNA interacts with the host cellular machinery to initiate protein synthesis. The VPg protein seems to play an important role in this early event, because removal of VPg protein with proteinase K from the genomic RNA abolishes RNA infectivity and, in addition, decreases its translation efficiency in vitro (Leonard et al., 2000; Herbert et al., 1997). The amino acid sequence of the potato virus Y genome-linked protein VPg corresponds to the amino acid residues 336 to 523 of the amino acid sequence identified under accession number CAA82642 in the GENBANK database (Nov. 14, 2006 version). It is reproduced herein as SEQ ID NO: 2.
Recent studies have reported an interaction between the VPg protein of certain potyviruses (turnip mosaic virus, lettuce mosaic virus, tobacco vein mottling virus, potato virus Y) and the plant translation initiation factor eIF4E (Wittmann et al., 1997; Duprat et al., 2002; Kang et al., 2005; Grzela et al., 2006). It has also been shown that the VPg protein encoded by certain vertebrate viruses of the Caliciviridae family, such as the feline calcivirus (FCV), the lordsdale virus (LDV)) or the recently discovered murine norovirus (MNV-1) interact with the cap-binding protein eIF4E (Goodfellow et al., 2005; Chaudhry et al., 2006).
Grzela et al. (2006) have found that the interaction between PVY VPg protein and eIF4E seems to have rather low species specificity since it was also observed with yeast as well as with the insect (S. frigiperda) eIF4Es. The authors have also described that the VPg/eIF4E interaction results in some inhibition of cell-free protein synthesis in insect cells, but without leading to insect cell death. However, the PVY VPg primary amino acid sequence does not exhibit the eIF4E-binding motif YxxxxLØ SEQ ID NO: 31) described above.